The general focus of this project is the role of calcium ions in the function of synapses between nerve cells. It is well-established that synaptic secretion of neurotransmitter chemicals serves as the communications link between nerve cells and that calcium ions directly trigger this secretion. It also is thought that calcium has another kind of action, one that regulates neurotransmitter secretion and produces certain forms of synaptic plasticity. The poorly-understood molecular mechanisms responsible for these actions of calcium are the subject of the proposed experiments. The specific goal of the project is to test the hypothesis that particular proteins found only on the membrane of synaptic vesicles - storage organelles that are involved in neurotransniitter secretion - mediate these actions of calcium. To test this hypothesis, chemicals probes that interfere with specific vesicle- associated proteins will be microinjected into living presynaptic terminals while assessing the ability of these probes to interfere with the triggering or regulation of neurotransmitter release. The hypothesis predicts that these chemical probes will interfere with neurotransmitter release only if the protein targets of the probes are involved in release. The particular vesicle-associated proteins to be considered include synaptotagmin, synaptobrevin, Ca/calmodulin-dependent protein kinase and Rab3a. Because few synapses have presynaptic terminals sufficiently large for this kind of microinjection experiment, the experiments will be performed on the unique 'giant' synapse of squid. The results of these experiments will help define the molecular mechanisms responsible for synaptic transmission and, thereby, ultimately will clarify the actions of the numerous neurological disorders that result from abnormal synaptic transmission.